Breast cancer is the most frequently diagnosed cancer among women in the United States with >200,000 new cases reported every year. Aggressive forms of breast cancers most commonly metastasize to bones, which constitutes the key factor for high morbidity rate and associated suffering. Metastatic breast tumor growth in bones leads to chronic pain in the limbs, pelvis and spines, which is undermanaged with existing pain medications, mainly morphine derivatives. As palliative care is the major therapeutic goal for patients with breast cancer bone metastasis, a better understanding of signaling crosstalk between metastatic bone tumor microenvironment and the adjacent sensory nerve fibers is necessary for the development of highly efficacious analgesics for chronic bone cancer pain. Metastatic breast cancer cells secrete high levels of parathyroid hormone-related peptide (PTHrP), which act on osteoblasts and osteoclasts to induce bone lysis/destruction and subsequent release of nutrients and cell signaling molecules that stimulate tumor growth. Here I propose a novel hypothesis that PTHrP, by acting through its receptor PTH1 on sensory afferents, induce constitutive nociceptor sensitization via upregulation of activity/expression of the transient receptor potential vanilloid-1 (TRPV1) channel, which might underlie a mechanism for chronic pain associated with metastatic breast/bone cancers. The TRPV1 channel is normally activated only at noxious temperatures (e43oC). However, PTHrP acting through the PTH1 receptor could induce phosphorylation of TRPV1 protein and lead to constitutive channel activation at body temperatures (d37oC), a mechanism that could underlie chronic pain in the absence of any overt stimulation. Aim 1 of my study will determine the specific cellular signaling events that underlie PTHrP-modulation of TRPV1 channel activity/expression in sensory neurons, and nociceptor sensitization. I will utilize patch-clamp electrophysiology, Ca2+ imaging, and membrane protein biochemistry to determine the molecular mechanisms underlying TRPV1 modulation by PTHrP. Aim 2 of my study will determine the role of PTHrP-modulation of TRPV1 in sensory afferents on chronic bone pain in vivo using scid mice xenografts of human breast cancer cells, MDA-MB231-BoM-1833, that metastasize to bones when injected intracardialy. I will use a series of un-evoked/spontaneous bone-related pain behavioral assessments in these mice with metastatic breast/bone tumors. I will also utilize pharmacological inhibition of PTHrP and TRPV1 in these mice, to further confirm the contribution of PTHrP-modulation of TRPV1 in chronic bone pain associated with metastatic breast/bone tumor growth. My proposed studies will advance our understanding of how the interplay between metastatic breast/bone tumors and sensory neurons mediate chronic pain. Findings from this study will significantly contribute to the development of effective pharmacotherapies for chronic pain associated with metastatic bone cancers.